The purpose of this study was to test the effect of repositioning, systematic displacements of the region of interest (ROI), and acquisition parameters (scan mode and integration time) on quantitative analysis of human trabecular bone microstructure at various skeletal sites, using microcomputed tomographic (microCT) technology. We investigated 28 cylindrical specimens of human trabecular bone (length 14 mm, diameter 8 mm) from four skeletal sites (femoral neck, greater trochanter, second lumbar vertebra, and distal radius). These specimens were selected from over 200 microCT measurements, in order to cover a large range of bone volume fraction (BV/TV) observed at each site. Cylindrical ROIs (length 6 mm, diameter 6 mm) were examined twice at an isotropic resolution of 26 microm, 8 weeks apart. In addition, comparative analyses were performed for displacements of the volumes of interest (VOIs) by 1, 2, 3, and 4 mm (83.4%, 66.6%, 50%, and 33.3% overlap), respectively. Eventually, comparative measurements were obtained at different resolution scan modes and integration times. The results show that microCT measurements are highly reproducible (range of the root mean square coefficient variation % (RMS CV%) = 0.64% to 1.29% for BV/TV at different sites). Displacements of the VOI of up to 4 mm generally led to non significant systematic differences in mean values of < 10%. When comparing various combinations of resolution scan modes and integration times, the use of an integration time of 100 ms was found to be preferable for determining microstructural parameters from human samples with this microCT scanner.